1. Field of the Invention
The present invention relates to a novel enzyme, and preparation and uses thereof. More particularly, the present invention relates to an enzyme which converts maltose into trehalose or converts trehalose into maltose (hereinafter designated as "maltose-trehalose converting enzyme"), as well as to preparation thereof. The present invention further relates to a microorganism capable of producing the enzyme, trehalose prepared with the enzyme, saccharide compositions containing the trehalose, and compositions containing the trehalose or the saccharide composition.
2. Description of the Prior Art
Trehalose or .alpha., .alpha.-trehalose has been known as a non-reducing saccharide consisting of glucoses. As is described in Advances in Carbohydrate Chemistry, Vol. 18, pp. 201-225 (1963), published by Academic Press, USA, and Applied and Environmental Microbiology, Vol. 56, pp. 3,213-3,215 (1990), trehalose widely exists in microorganisms, mushrooms, insects, etc., though the content is relatively low. Trehalose is a non-reducing saccharide, so that it neither reacts with substances containing amino groups such as amino acids and proteins, induces the amino-carbonyl reaction, nor deteriorates amino acid-containing substances. Thus, trehalose can be used without a fear of causing an unsatisfactory browning and deterioration. Because of these, the establishment of an industrial-scale preparation of trehalose has been in great demand.
Conventional preparations of trehalose are, for example, those disclosed in Japanese Patent Laid-Open No. 154,485/75 wherein microorganisms are utilized, and in Japanese Patent Laid-Open No. 216,695/83 wherein maltose is converted into trehalose by the combination use of maltose- and trehalose-phosphorylases. The former, however, is not suitable for the industrial-scale preparation because the content of trehalose present in microorganisms as a starting material is usually lower than 15 w/w % (the wording "w/w %" is abbreviated as "%" in the specification, unless otherwise specified), on a dry solid basis (d.s.b.), and the extraction and purification steps are complicated. The latter has the following demerits: (i) Since trehalose is formed via glucose-1-phosphate, the concentration of maltose as a substrate could not be set to a satisfactorily high-level; (ii) the enzymatic reaction systems of the phosphorylases are reversible reactions, and their yields of the objective trehalose are relatively low; and (iii) it is substantially difficult to retain their reaction systems stably and to proceed their enzymatic reactions smoothly. Therefore, the aforesaid conventional preparations could not be used as an industrial-scale preparation.
It is known that partial starch hydrolysates, prepared from a material starch such as liquefied starch, dextrins and maltooligosaccharides, usually exhibit a reducing power because of their reducing end groups. The reducing power is generally expressed by "Dextrose Equivalent (DE) value" based on the dry weight. It is known that among reducing partial starch hydrolysates those with a relatively-high DE value generally have a considerably-low molecular weight and viscosity, as well as a relatively-high level of sweetness and reactivity, and readily react with substances having amino groups such as amino acids and proteins to cause an unsatisfactory browning, smell and deterioration of their quality. Since the properties of reducing partial starch hydrolysates are varied dependently on their DE values, the relationship between reducing partial starch hydrolysates and their DE values is significant. It has been even believed impossible to break away the relationship in this field.
As regards the preparation of trehalose, it is reported in the column titled "Oligosaccharides" in the chapter of "Current Status of Starch Application Development and Related Problems" in "Food Chemicals", No. 88, pp. 67-72 (August, 1992) that "In spite of a wide applicability of trehalose, the enzymatic preparation via a direct saccharide-transfer reaction or a hydrolytic reaction has been reported to be scientifically almost impossible in this field." Thus, the preparation of trehalose by an enzymatic reaction using starch as a material has been deemed scientifically very difficult.
The present inventors, however, had changed this common sense and succeeded to establish a preparation of trehalose as disclosed in Japanese Patent Application No. 362,131/92 wherein trehalose is directly produced from non-reducing partial starch hydrolysates by allowing glucoamylase together with a non-reducing saccharide-forming enzyme capable of forming non-reducing saccharides, having a trehalose structure as an end unit and having a degree of glucose polymerization of 3 or higher, to act on reducing partial starch hydrolysates having a degree of glucose polymerization of 3 or higher, prepared from a material starch. The method, however, requires 2 or more types of enzymes and employs as a material a relatively-high molecular weight amylaceous saccharide having a degree of glucose polymerization of 3 or higher as well as a relatively-high viscosity. In addition, the saccharide composition of the resultant product is considerably complicated, and this may result in a high production cost. Therefore, the establishment of a novel preparation of trehalose in which trehalose is formed from maltose and partial starch hydrolysates having a degree of glucose polymerization of 2, both of which are industrially produced, stably supplied and commercially available.